Microfluidic devices have attracted great interest as they to provide a platform for performing analyses on extremely small volumes of fluid, and when produced utilizing photolithography techniques can be manufactured inexpensively. These devices have the potential to act as a “lab on a chip”, integrating multiple functionalities including, for example, sample preparation, thermal cycling to support the polymerase chain reaction, and absorbance or fluorescence monitoring. Their compact size makes them particularly suitable for use in portable devices, potentially allowing the performance of sophisticated analyses in a clinician's office or in the field. One of the challenges with using microfluidic devices in the analysis of multiple samples, however, is sample compartmentalization. While a conventional laboratory analyzer may utilize a series of cuvettes or similar receptacles to prevent contamination between samples this approach is difficult to implement with small volumes of fluid, where interactions with device surfaces can supersede bulk flow properties.
Typical microfluidic devices utilize a single fluid phase that flows continuously through the device. Introduction of a discrete volume of fluid test sample or reagent into such a device leads to the formation of a fluid segment that moves through the channels of the apparatus. Unfortunately, such a fluid segment will tend to become dispersed due to forces such as diffusion and turbulence within the flow channel. In addition it is possible for components of the fluid segment to interact with the walls of channels of the microfluidic device, only to be released at a later time. Such phenomena can result in contamination between fluid segments and results in the need to design such microfluidic chips with features to reduce turbulence within fluid channels and to design test protocols that incorporate time consuming washing or flushing of the interior volume between samples. In addition, dispersion of fluid segments makes it difficult to provide reproducible volumes and concentrations of fluid segment contents for characterization reactions.
One approach to resolving this issue has been the introduction of digital microfluidic devices, in which sample fluids for analysis or other treatment are introduced into the channels of the device in the form of discrete, low volume droplets. For example, introducing aqueous samples with biochemical or biological contents as aqueous droplets that travel within a channel containing an immiscible oil medium reduces interaction with the channel wall and prevents dispersion, minimizing contamination between droplets. Reagents used in the characterization of such samples can be treated similarly. In order to be effective, however, a digital microfluidic device requires a mechanism for high-speed droplet generation with precise volume control in order to fully realize accurate, high throughput analysis.
Passive mechanisms may be used for rapid, continuous droplet generation as a function of flow through such a device. Highly uniform droplets can be generated at a rate of thousands of drops per second in this fashion (Yobas et al. (2006) Lab on a Chip, 6:1073-1079). U.S. Pat. No. 7,759,111 describes such a device, where droplets are sheared from a stream of aqueous media by a flow of immiscible oil. Another example of a passive device is disclosed in WO 2010/110843A1, in which a barrier intruding into a fluid channel acts in combination with fluid and flow characteristics of the channel to form vortices that provide periodic variations in pressure that drive droplet formation. Such devices, however, do not provide on demand generation of a droplet containing specifically designated volume of sample fluid (for example, a volume containing a particular cell of interest) and do not lend themselves to the production of individual droplets with different volumes. This limits their utility for the characterization of different samples volumes and in the performance of a variety of testing protocols.
Active methods for droplet generation, which rely on the use of an applied force to drive droplet formation, can address these issues. Such devices may incorporate physical components that regulate flow through the device. One example of this is the use of pneumatically driven microvalves that are integrated into the microfluidic device (Zeng et al. (2009) Lab on a Chip 9:134-1343), which permitted controlled droplet formation at rates as high as 100 droplets per second. Another example of this approach is the use of a movable wall of flexible material (PDMS) that is integrated into the microfluidic chip and driven by air pressure to periodically interrupt the flow of a fluid phase in order to provide a dispersion (Hsiung et al. (2006) J. Micromechanics and Microengineering, 16: 2403-2410), which demonstrated rates of droplet formation as high as 20 per second. Yet another example, US 2010/0059120, discloses the use of a two channels connected by an opening, in which a flow interruptor in one channel can be triggered to block fluid flow and force a portion of its contents into the second channel. Another example of such a device is described in US 2010/0163412, which discloses a device that incorporates a flexible fluid reservoir that is compressed briefly by an imbedded piezoelectric device to generate pressure for droplet formation. Such features add significantly complexity to the design of these microfluidic devices, further complicating the manufacturing process. The mechanical nature of such approaches limits the frequency at which droplets can be produced and may show changes in performance over time. In addition, these approaches tend to produces droplet populations with greater variation in droplet size than those produced using passive devices.
Other approaches to active droplet generation have relied on the use of massless or essentially massless energies applied to the device in order to avoid the disadvantages of mechanical components. Some of these utilize the application of electrical fields to the device to alter fluid flow or change the properties of the interface between two fluids in order to facilitate droplet formation. This can require large differences in conductivity between the fluids involved, which limits the utility of such devices. For example, US 2006/0231398 discloses the use of potential differences to move droplets between immiscible low and high resistance fluids by electrowetting, utilizing a potential difference to temporarily lower the surface tension at the interface between the fluids until the existing flow pattern is sufficient to generate droplets. A similar approach is described in WO 2010/151776, in which a potential difference drives a combination of effects, including electrokinetic flow and interference in the interface between two immiscible fluids, to generate droplets. Yet another example of the use of potential differences to drive droplet formation is found in WO 2011/023405A1, which discloses a combination of a nozzle structure and establishment of a potential difference to electrospray droplets of a conductive fluid into a fluid dielectric. An approach that does not require large conductivity differences between the fluids involved in droplet formation is disclosed in US 2005/0031657, which describes heating a portion of a container within the device using a resistance heater until a portion of the fluid stored therein is vaporized. Pressure from the vaporized fluid pushes a portion of remaining fluid through a nozzle into an immiscible fluid. Droplet generation from this approach is relatively slow, however, producing only around 15-25 droplets per second per nozzle. While these approaches avoid the use of mechanical components, they require the incorporation of electrodes, resistance heaters, or similar components into the device. This adds complexity to the design of the device and further requires the use of supporting features for reliably supplying current.